Analytic device including nanostructures

ABSTRACT

A device for detecting an analyte in a sample comprising: an array including a plurality of pixels, each pixel including a nanochain comprising: a first nanostructure, a second nanostructure, and a third nanostructure, wherein size of the first nanostructure is larger than that of the second nanostructure, and size of the second nanostructure is larger than that of the third nanostructure, and wherein the first nanostructure, the second nanostructure, and the third nanostructure are positioned on a substrate such that when the nanochain is excited by an energy, an optical field between the second nanostructure and the third nanostructure is stronger than an optical field between the first nanostructure and the second nanostructure, wherein the array is configured to receive a sample; and a detector arranged to collect spectral data from a plurality of pixels of the array.

PRIORITY CLAIM

This application claims the benefit under 35 USC 371 to InternationalApplication No. PCT/IB2014/003194, filed Dec. 23, 2014, which claimspriority to U.S. Provisional Patent Application No. 61/920,725, filedDec. 24, 2013, each of which is incorporated by reference in itsentirety.

TECHNICAL FIELD

The invention relates to a device that uses nanostructures to analyzesamples.

BACKGROUND

Raman spectroscopy is a technique that utilizes Raman scattering bymolecules to assess the molecules and allow the molecules to beidentified by spectral signature patterns.

SUMMARY

In one aspect, a device for detecting an analyte in a sample can includean array including a plurality of pixels, each pixel including ananochain comprising a first nanostructure, a second nanostructure, anda third nanostructure, wherein size of the first nanostructure is largerthan that of the second nanostructure, and size of the secondnanostructure is larger than that of the third nanostructure, andwherein the first nanostructure, the second nanostructure, and the thirdnanostructure are positioned on a substrate such that when the nanochainis excited by an energy, an optical field between the secondnanostructure and the third nanostructure is stronger than an opticalfield between the first nanostructure and the second nanostructure,wherein the array can be configured to receive a sample; and a detectorarranged to collect spectral data from a plurality of pixels of thearray.

In certain embodiments, the first nanostructure can include a metal, thesecond nanostructure can include a metal, and the third nanostructurecan include a metal. The nanostructure can include silver, gold,platinum, or aluminum.

In certain embodiments, the second nanostructure can be positionedbetween the first nanostructure and the third nanostructure. Thedistance between the first nanostructure and the second nanostructurecan be less than the diameter of the second nanostructure. The distancebetween the second nanostructure and the third nanostructure can be lessthan the diameter of the third nanostructure.

The device can include 10 by 10 pixels. The distance between two pixelscan be at least 1 μm. Each pixel can deliver spectral data for at leastone analyte. Each pixel can be capable of detecting at least twodifferent kinds of analytes. The device can be capable of detecting atleast 5 different kinds of analytes. The device can be capable ofdetecting at least 10 different kinds of analytes. The device can becapable of detecting at least 20 different kinds of analytes.

In certain embodiments, the analyte can be detectable by Raman spectrum.The analyte can be a peptide. The device can be capable of detecting asingle molecule.

In certain embodiments, the first nanostructure is a nanosphere, thesecond nanostructure is a nanosphere, and the third nanostructure is ananosphere. The diameter of the first nanostructure can be less than 500nm. The diameter of the second nanostructure can be less than 200 nm.The diameter of the second nanostructure can be less than 100 nm.

In another aspect, a method for detecting an analyte can includecontacting an analyte with an array including a plurality of pixels,each pixel including a nanochain comprising a first nanostructure, asecond nanostructure, and a third nanostructure, wherein size of thefirst nanostructure is larger than that of the second nanostructure, andsize of the second nanostructure is larger than that of the thirdnanostructure, and wherein the first nanostructure, the secondnanostructure, and the third nanostructure are positioned on a substratesuch that when the nanochain is excited by an energy, an optical fieldbetween the second nanostructure and the third nanostructure is strongerthan the optical field between the first nanostructure and the secondnanostructure; and collecting spectral data from a plurality of pixelsof the array.

In certain embodiments, the method can include taking Raman spectrum ofthe nanochain. The analyte can include a peptide. The firstnanostructure can include a metal, the second nanostructure can includea metal, and the third nanostructure can include a metal. Thenanostructure can include silver, gold, platinum, or aluminum. Incertain embodiments, the second nanostructure can be positioned betweenthe first nanostructure and the third nanostructure.

In certain embodiments, the method can detect a single molecule.

Other aspects, embodiments, and features will be apparent from thefollowing description, the drawings, and the claims.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1A shows an SEM image of a patterned area and FIG. 1B shows thenanochain after electroless growth.

FIGS. 2A-2D show a schematic representation of a nanochain formation.

FIG. 3A shows an optical image of a scanned area on a device containinga nanochain; FIG. 3B shows spectra measured at different positions ofthe device; FIG. 3C shows a mapping analysis of the intensity Raman peakat specific wavenumber a649 cm⁻¹.

FIG. 4 shows FDTD numerical simulation of a single nanochain.

FIG. 5 shows mapping analysis of a nanochain including threenanostructures.

FIGS. 6A-6D show mapping analysis of different nanochain structures(monomer, dimer, trimer, tetramer).

FIG. 7A shows mapping analysis for a nanochain with three nanospheres;FIG. 7B shows mapping analysis for a nanochain with two nanospheres.

FIGS. 8A-8B show SEM images of nanostructures with different sizes.

FIGS. 9A-9B show SEM images of nanostructures with different sizes.

FIGS. 10A-10C shows SEM images of nanostructures with different sizesand numbers on nanochains (monomer, dimer, trimer tetramer).

FIG. 11 shows an SEM image of a device including a plurality of pixels.

FIG. 12 shows SEM images of a device including a plurality of pixels andSEM images of three different pixels typology (dimers, trimers,tetramers).

FIGS. 13A-13B show a wild type and a mutated peptide W1837 from geneBRCA1 in breast cancer.

FIGS. 14A-14B shows Raman spectra of W1837 peptide on a device includingnanochains.

FIGS. 15A-15C show Raman spectra of a specific pixel from a mixture of22 peptides on a device including nanochains.

DETAILED DESCRIPTION

Nano-optics system offers a great possibility to control the propagationof light at the surface of nanometals due to the fact that nanometallicstructure overcomes the diffraction limit, observed in the conventionaloptics.

A device for detecting an analyte can include a nanochain comprising ofthree nanostructures. The device can also include a plurality of pixels,where each pixel includes at least one nanochain. A nanochain caninclude a first nanostructure, a second nanostructure, and a thirdnanostructure. The nanostructures can be aligned with decreasing size.For example, size of the first nanostructure can be larger than that ofthe second nanostructure, and size of the second nanostructure can belarger than that of the third nanostructure. The second nanostructurecan be positioned between the first nanostructure and the thirdnanostructure.

The nanostructures can be a metal. At visible regime, a noble metal canbe preferred, such as a silver or a gold, or others. In the ultra violetregime, the metal can be a noble metal, such as silver, and the metalcan be other type of metal, such as aluminum. When a nanochain isexcited by an energy, an optical field or an electric field between thesecond nanostructure and the third nanostructure can be stronger thanthe field between the first nanostructure and the second nanostructure.

The device can include any number of pixels, for example 10×10 pixels.The distance between the pixels can be controlled such that the devicecan provide a detection signal for only one pixel. In one example, thedistance between the pixels can be at least 1 μm. The distance betweenthe nanostructures can also be controlled with precision of nanometers.

The device can detect an analyte by Raman and/or fluorescencespectroscopy. The analyte can include a peptide. The device can detect amixture including more than one type of analyte, such as 20 peptides.The device can also detect the existence or absence of a singlemolecule.

Surface Enhanced Raman Spectroscopy (SERS)

The intensity of a Raman spectrum of a molecule that is in closeproximity to an appropriate noble metal surface can be remarkably largerthan in the absence of the surface. This phenomenon can be due to lightexcitation of charge density fluctuations on the metallic surface, whichin turn creates a strong electromagnetic field nearby. The frequency ofthe radiated field can be close to that of the incident light, while,spatially, the field can be characterized by strongly localized regionsof low and high intensity (higher than the intensity of the incidentlight). A molecule can emit Raman scattered light with a magnitudeproportional to the fourth power of the local value of the field.

The overall enhancement in Raman signal from a molecule located in ahigh intensity region near the surface can exceed eight orders ofmagnitude. Furthermore, the strong localization of the enhancementregions makes it possible to resolve Raman signal from objects of lengthscales comparable to that of these regions.

The field distribution that radiates from a surface can depend on thegeometry of the SERS device. In one example, nanostructures can beformed by aligning three metal nanostructures, such as silvernanospheres, of decreasing diameter. A dramatic increase in fieldintensity can be obtained in the gap between the two smallernanostructures. The gap can be 5-20 nm wide. A device with such analignment can have a tunable spectral maximum for the enhancement.

Preparation of a Nanochain

The progress in plasmonics device architecture has also been supportedby the advancement in the nanofabrication techniques. With the fastdevelopment of plasmonics area and the application of nanoplasmonicdevice, from physics to biology and engineering to medical field, it canbe important to fabricate a device that demonstrates the generation ofhigh electromagnetic field to analyze a substance, for example, usingspectroscopic techniques. The device reproducibility is important aswell.

There are many routes to fabricate SERS devices, such as metal islandfilms, such as electrochemically modified electrodes, nanospherelithography, electron beam lithography, etc. See, for example,Costantino et al, Anal. Chem. 73, 2001, 3674-3678; D L Jeanmaire et al.,J. Electroanal. Chem. 84, 1977, 1, each of which is incorporated byreference in its entirety. One way is to produce Ag colloids and thenattach these metal colloids to biomolecules. See, for example, K. Kneippet al, Phys. Rev. Letts. 78, 1997, 1667, which is incorporated byreference in its entirety. Continuous spectral fluctuations in Ramanintensity can be observed at different times for SERS substrates. Inother terms, SERS substrate prepared by utilizing colloidal techniquecan show statistical signal variations. Efforts are being made to theperiodic reproducible and controllable SERS substrates, which have beenutilized to carry out the analysis for various molecules (Rhodamine6G,protein, etc.). See, for example, G. Das et al., Biosensors andBioelectronics, 24, 2009, 1693-1699; F. De Angelis et al., Nano Letts.8(8), 2008, 2321-2327, each of which is incorporated by reference in itsentirety. Another kind of SERS device can be fabricated. See, forexample, K. Li et al., Phys. Rev. Letts. 91 (2003) 227402; J. Dai, etal., Phys. Rev. B 77 (2008) 115419, each of which is incorporated byreference in its entirety.

A nanochain includes at least three nanostructures. A nanostructure is astructure with a size of less than 1 μm. A nanostructure can be ananolense, a nanosphere, and so on. In one example, a nanochain caninclude three nanostructures aligned with decreasing size. A nanochaincan be prepared via the combination of high precision electron beamlithography and electroless plating.

Electroless growth is a process where metal ions are reduced anddeposited as metals upon a pre-prepared silicon surface. Metal grainsmay be accordingly obtained with an average size as small as fewnanometers, with an overall shape and dimension that can be regulatedwith a high precision and reproducibility.

Building the nanostructures can include several steps requiring highperformance fabrication technologies. First, the nanostructure patterncan be fabricated in a film of polymethylmethacrylate (PMMA-A2) polymer(the resist) coated onto a silicon surface. The pattern can includethree aligned cylindrical cavities of different diameters; the firstcavity can have a diameter of between 500 to 100 nm; the second cavitycan have a diameter of between 200 and 50 nm; the third cavity can havea diameter of between 100 and 5 nm. The height of the cavities can bebetween 20 and 150 nm. These cavities can be at different distances. Forexample, the distance between the first cavity and the second cavity canbe between 10-100 nm, and the distance between the second cavity and thethird cavity can be 5-50 nm.

The pattern can be realized using an ultra-high-resolution electron-beamlithography (EBL) technique. In EBL, a highly focused electron-beam isused to depolymerize regions of the resist of controlled geometry. Thedepolymerized part of the resist can be then developed by immersion inan appropriate solvent and a mold for the successive deposition of ametal, such as a silver, is obtained. The features of a mold used forthis can be controlled. After patterning with EBL and rinsing, a highlyclean silicon surface can be obtained inside cavities.

This process can include immersing the nanostructure pattern in a watersolution containing a metal salt, such as AgNO₃, and an acid, such asHF. The acid can react with the SiO₂ on the surface of the exposedsilicon at the bottom of the cavities and initiates a series of stepsthat eventually leads to transferring electrons from the solution to thesilicon. The metal ions from the dissociated salt that diffuse to thesurface can be reduced by these electrons to Ag which is adsorbed on thesurface via ionic interactions. The metallic Ag structures that grow viathese steps can act as an electrode charged by the electrons in thesilicon and further contribute to the reduction of silver ions from thesolution. Since no external electrical field is required to drive thesereactions, this type of deposition process is usually referred to aselectroless plating. After the deposition of metal particles, the samplecan be rinsed with bi-distilled water to wash out the salt from thenanostructure's surface.

Optical properties of nanoparticles, such as metallic nanoparticles, andtheir aggregates can be interesting. Such nanoparticles can induce localoptical fields that exceed the exciting fields. Raman scattering can beincreased, which allows for the detection and spectroscopy of singlemolecules. The Raman enhancement can be proportional to the fourth powerof the local field. Enhanced Raman scattering can exist at the gapsbetween metal nanostructures.

A Device Including a Nanochain

Nanostructure plasmonic devices, based on nanoholes, nanoantenna,nanocolloids, metal nanoparticles (in particular, gold, silver, orplatinum) of varying shape, size, size distribution, etc., have beenproposed to enhance the optical signals of interested molecules,attached or located in close proximity to nanometer-sized metallicstructure. See, for example, M. Moskovits, Rev. Mod. Phys. 57, 1985,783; J J Mock, et al., J. Chemical Physics, 116, 2002, 6755, each ofwhich is incorporated by reference in its entirety. When laser radiationinteracts with sharp metallic surfaces, the increase in localizedelectric field can induce an enhancement in the Raman signal. Theincrease in Raman scattering may be large. See, for example, K. Kneipp,et al., Phys. Rev. Letts. 78, 1997, 1667; S. Nie, et al., Science 275,1997, 1102, each of which is incorporated by reference in its entirety.The increase delivers the possibility of detecting very little amount ofmolecules, at the limit of single molecule situated in the proximity ofthe metallic surface.

For metal having resonant quality factor ‘Q’ and number of nanosphere‘n’, the theoretical effective SERS enhancement G_(SERS) can be Q^(4n).When considering Ag-based nanochains (Q˜30) constituted by threenanospheres, the theorectical SERS enhancement factor can be in theorder of 10¹² or more. An SERS device can be made of a self-similarchain. A self-similar chain can be a chain including nanospheres wherethe ratios between the subsequent radiuses and gaps are constant. Ananochain can include three metal nanospheres. Two techniques (e-beamlithography technique and metal electroless deposition) can be combinedto fabricate this device. The use of well-defined nanolithographicstructures allows a better control of the SERS substrate, giving thepossibility to fabricate a controllable and reproducible device. Tosatisfy the inflexible nanostructure size and shape, electron beamlithography technique can be taken into consideration. Whereas, thesite-selective silver nanoparticle using electroless depositiontechnique can be utilized for augmenting the electromagnetic field.Theoretical simulation using Finite Difference Time Domain (FDTD) can beperformed for the self-similar nanochain SERS device, keeping the realcharacteristic parameters; dielectric constant of the polymer, Q factorof silver, the experimentally observed the size of the nanospheres andthe distance between the two consecutive silver nanospheres, etc. and isfound to have enhancement factor about 10⁷-10⁸. This signal enhancementvalue is close to what it is observed by self-similar nanochains SERSsubstrate fabricated after depositing various molecules on it.

Nanochain SERS biophotonics device can be designed and fabricated. Thedevice exhibits the few molecules detection capabilities for Rhodamine6G (R6G), site selective deposited carbon, DNA, control and mutated BRCApeptide. In this nano-optics system, electro-magnetic light confinesefficiently in the gap between the two smallest Ag nanospheres due tolocalized surface plasmon in such a manner that it leads to ananolocalization of e.-m. field and so the Raman signal intensity.

An analytical device can include a nanochain comprising nanostructures.The nanostructures are metallic. The material that can be used includessilver, gold, aluminum. The nanostructures can be in an array of three,aligned with decreasing size. The surface and shape of nanostructurescan be disordered (the nominal center of gravity of each nanostructurecan be aligned with an accuracy better than 2 nm). The analytical devicecan be used to detect an analyte. The analytical device can be used forgeneric molecules and in particular for protein detection from a sample,such as a human sample. The device can include a plurality of pixels.The pixels can form an array. Each pixel can be the sensitive unit ofthe device. The device can produce a strong localization of electricalfield when the nanochain is illuminated by a laser light. Various laserwavelength regimes, such as visible wavelength regime, can be used. Ananochain can include three nanospheres. The localization of theelectrical field can be mainly between the smallest sphere and themiddle sphere. In a three-nanostructure chain aligned with decreasingsize, the electrical field between the smallest nanostructure and themid-sized nanostructure can be stronger than that between the largestnanostructure and the mid-sized nanostructure.

A nanochain can generate a localized electrical field when illuminatedby a laser beam. If molecules are deposited on the nanochain, theyscatter the laser light that can be analysed and detected by a Ramanspectroscope apparatus. The information obtained by this apparatus canbe a specific signature of the molecule that entered the sensitive areaof the nanochain.

A nanochain array can be fabricated and used to analyse a material. Thematerial can be a mixture including at least one type of analyte. Forexample, a nanochain array can detect a mixture of about 20 peptidesfrom a human sample of breast cancer. The mixture can be resolvedcomponent by component.

The footprint of the electrical field can cover a region, whose size canbe about 5×5 nm². This indicates that if a solute is deposited on ananochain, only the molecules that enter this region can be detected.The device may not be able to detect molecules outside of this region.The molecules can be detected through a spectroscopy, such as a Ramanspectroscopy.

The device can detect and analyze composition of highly diluted mixturesof molecules, such as in the range of nanomolar. This can be applied toearly cancer diagnosis or pollutant environmental analysis and others.The device can be used when there is a mixture of soluble molecules in ageneric solvent and the detection can be combined with a spectroscopytechnique such as Raman or Fluorescence. The sample can be a dilutedsample. The dilution factor can be such that the concentration of ananalyte of interest in the sample leads to a significant probabilitythat one molecule can be found at a “hot spot” (the sensitive region) ofthe nanolens. The measured experimental deposition rate in the hot spotis between 0.25 to 1%. This means that even if 99% of molecules gooutside the hot spot, the sensitivity of the nanochain allows thedetection. This does not require sophisticated means or tools to depositthe molecules exclusively in the hot spot area. For example, an analyteconcentration can be between 1 and 100 nanomolar, for example between 2nanomolar and 25 nanomolar. This can be a range used with a deviceincluding a nanochain.

The further improvement in the functioning of the device can be due tothe intrinsic disorder or roughness of a surface of each nanosphere.This disorder can contribute to a larger localization of the electricalfield.

The working principle of the device can be due to plasmonics andnano-localization of electrical field for obtaining a strong Ramansignal even with few molecules in the pixel. It can further exploit thenatural disorder arising in the fabrication of the nanospheres togreatly enhance the electric field and achieve a high sensitivity. Thescale of sensitive area can be strongly localized. The device can solvehighly diluted mixture by physical means, label free with no additionalchemical reactions to detect the composition of the solute. The designand the use of the array can solve a mixture of peptides that bring thesignature of a tumor. The reconstruction of mixture composition can berealized by reading the Raman spectra of each pixel in the matrix array.With proper dilution, each pixel can contain, randomly distributed, onlypart of the mixture component. Reading all pixels, all components of themixture can be recovered. Typically in each pixel, between 2 to 4components dominate. These are weighted by a linear combination of purecomponents. The operation is repeated on each pixel.

The advantages of the device include that it is sensitive to lowconcentration of solute down to nanomolar, it gives a lot of informationon the chemical state of the molecules composing the mixture, such as awild type or a mutated form of peptides, and the quantity of solute canbe very low, such as below the nanogram range.

Examples

The device fabrication can involve several steps requiring highperformance fabrication technologies. The nanochain pattern can bewritten using an ultra-high-resolution e-beam lithography (EBL)technique, realized on a thin layer of PMMA-A2 resist (50 nm), which isspun onto a cleaned silicon substrate. The stability of the beam and itssettings (45 pA of beam current at 50 keV acceleration voltage, beamsize—2 nm) generated by EBL system (CRESTEC CABL-9000C), the features ofthe resist used, and the development process (1 min rinsing with IPA at4° C.) help produce a controllable nanostructure. The dimension of thepatterned SERS device is around 320 nm and the two small gaps betweenthe two consecutive nanospheres are 20.6 nm and 18.5 nm, respectively,as shown in FIG. 1A). The smallest nanohole on resist is of 33.6 nmdiameter. Various kinds of nanochains structures with single, double,quadruple lenses are fabricated in order to test and compare thecapability to localize the Plasmon Polaritons (PPs) close to thesmallest nanospheres.

The strategy to perform a highly controlled nanochemistry over thepost-lithographic surface was an important step to make a site-selectivemetal deposition. The electroless process can be used to depositAg-metal on a predefined area in which the reduction of oxidizedmetallic species led to formation of neutral metal atom. See, forexample, D. V. Goia, et al., New J. Chem. 22, 1998, 1203, which isincorporated by reference in its entirety. Ag-nanometal deposition canbe accomplished using the solution of AgNO₃ and HF. See, for example, S.Yae et al, Electrichimica Acta 53, 2007, 35, which is incorporated byreference in its entirety. Right after the deposition of metalparticles, the sample was rinsed with bidistilled water to wash out thesalt from the nanostructure surface. After the Ag-electrolessdeposition, the two small gaps between the lenses are 37.7 nm and 8.85nm, whereas the diameter of smallest nanosphere becomes 25.8 nm, shownin FIG. 1B. Nanostructure devices with two, three and four nanochainscan also be prepared, as shown in FIGS. 10A-10C. A pixel of a chip caninclude 2 nanochains (a dimer), 3 nanochains (a trimer), or 4 nanochains(a tetramer). The fabrication process in summary is also illustrated inFIGS. 2A-2D. After the resist spin coating (FIG. 2A), the lithographyprocess creates nanohole (FIG. 2B), which can be further used for metalsite-selective deposition. The desposition in the nanohole (FIG. 2C)ultimately forms the self-similar nanochain substrate (FIG. 2D).Au-based nano-optics devices can also be fabricated following the sametechnique.

The device can be used as an SERS device. Micro-Raman spectra (inViaRaman Microscope, Renishaw) of the device were excited by using 514 nmAr+ laser (Power=0.018 mW and accumulation time=4 sec) in backscatteringgeometry with the spectral resolution of about 1.3 cm⁻¹. Highmagnification objective 150× is used for all the Raman measurements. XYRaman mapping measurements were carried out with the minimum step sizeof 100 nm.

R6G (an organic fluorescent molecule) monolayer was deposited over thedevice using immersion technique and thereafter it was rinsed bybidistilled water to wash out the excess molecules not depositeddirectly on the metal surface. Micro-Raman measurements are performed inthe area that consists of one nanochain structure, shown in FIG. 3A. Twostatic Raman spectra, one over nanostructure (top line) and the otherone outside the nanochain (bottom line), are performed in the range of1050-1925 cm⁻¹ (FIG. 3b ).

The static measurement points can be seen in optical image of themapping area. R6G SERS peaks are observed at around 1360 cm⁻¹ attributedto COO⁻ vibration; 1506, 1569, 1649 cm⁻¹ related to the C—C ringstretching vibration. Besides these bands, a few more bands, centered at1126 and 1188 cm-1 attributed to C—H_(x) in-plane bending, and 1310 cm⁻¹attributed to C—OH stretching vibrations, are also evident. It can beobserved from FIG. 3B that the Raman spectra of R6G are observed only atthe nanostructure point, whereas only a broad background is observed atthe rest of the location. Since the molecule deposited on theAg-nanoparticles is a fluorescent molecule which, in general, shows abroad fluorescence background spectrum, the presence of a nanochaincauses a fluorescent quenching and hence only the Raman spectrum of themolecule was shown. This phenomenon of fluorescent quenching for themolecules attached to the metal surface has been observed. See, forexample, E. Dulkeith et al., Phys. Rev. Letts., 89 (2002) 203002-1; P.Anger et al., Phys. Rev. Letts., 96 (2006) 113002-1, each of which isincorporated by reference in its entirety. Micro-Raman XY-mappingmeasurements were carried out in the range of 1050-1925 cm⁻¹ for thearea of around 3×2 μm². The mapping analysis, keeping the R6G peakcentered at 1649 cm⁻¹ as a reference band, is shown in FIG. 3C. Thepolarization of incident source is parallel to the nanostructure whichis an optimal condition to achieve the maximum SERS signal. The Ramananalysis has been executed by selecting the Raman band in the region of1618 and 1689 cm⁻¹. For the mapping analysis, each and every spectra inthe region 1618-1689 cm-1 was first baseline corrected and thenevaluated after the intensity of the band was centered at 1649 cm-1. Themaximum peak intensity is observed only at those points where nanochainstructures are present. The remaining area doesn't show any variationregarding this vibrational band. High sensitive, and highsignal-to-noise SERS signal are observed for R6G using this device. Theobserved broadness of the mapping band is due to the convolution oflaser beam-width. Considering that the major SERS contribution is fromsmallest metal nanosphere and the surface area of a Rhodamine6G moleculeis about 0.9 nm², the Raman signal coming from the nanochain can be dueto closely packed 100-150 molecules.

If there are metallic nanospheres in self-similar cascade mode, themajority of SERS intensity can come from the vicinity of the smallestnanosphere and the enhancement factor can reach >10¹². Fine DifferenceTime Domain (FDTD) simulation can be performed and then compared betweenthe real geometry device fabricated in lab and the ideal device proposedby Dai. See, for example, J. Dai et al., Phys. Rev. B 77 (2008) 115419,which is incorporated by reference in its entirety. The device inself-similar mode keeps functioning even when the device geometry isdeformed. In addition, The Stockman used 380 nm laser wavelength whichis the best for the silver nanoparticles to simulate the fieldenhancement whereas 514 nm was used as a SERS spectra excitationwavelength. Considering that the major SERS contribution is fromsmallest metal nanosphere and the surface area of a Rhodamine6G moleculeis about 0.9 nm², it is estimated that the Raman signal coming from thenanochain is due to 100-150 molecules when closely packed. Taking intoconsideration all the factors (such as the number of molecules observedunder laser spot, laser power, accumulation time, Raman intensity), theenhancement factor using the nanochain device is calculated to be around5×10⁷ with respect to the Si.

The experimental verification of high sensitivity by varying theposition of molecules on self-similar chain device is carried out bydepositing the carbon at a known position of the nanochain device. Afterdepositing the carbon on the top of smallest, middle and biggestnanosphere, individually, the micro-Raman measurements were performed.3D-Raman intensity analysis related to the C—C vibration centred at 1561cm-1 is performed for each and every spectrum in the mapping area byselecting the base line corrected curve in the region of 1400-1750 cm-1.It can be noticed that the band intensity is maximum for the sample onwhich carbon is deposited on the smallest nanosphere. The intensitytrend of I_(smallest)>I_(middle)>I_(biggest) is observed. Thesemeasurements confirm the detection sensitivity of this device. See FIGS.4 and 5.

The measurements are performed to verify the dependency of Raman signalby increasing the number of nanochains, shown in FIGS. 6A-6D and FIGS.10A-10C. R6G monolayer is deposited over the device and the Ramanspectra were taken with the same Raman parameters. The mapping analysisresults, shown in FIGS. 6A-6D, is performed for the Raman band centeredat 1649 cm⁻¹. In the inset, the associated nanochain structure is shown.3D mapping spectra in FIGS. 6A-6D illustrate the increase in Ramansignal from 700 for single nanochain to around 2000 counts forfour-nanochain structure.

Raman intensity for nanochains with three nanospheres and twonanospheres was compared. The results show the validity of a deviceincluding three nanospheres in cascade mode. For such verification,monolayer benzenethiol molecule is deposited over a device with twonanospheres and three nanospheres. Micro-Raman mapping analysis for theRaman band centred at 1584 cm⁻¹ is carried out for both samples, shownin FIGS. 7A-7B. The peak intensity increase and signal to noise ratioimprovement were observed for the device with three nanospheres.

To prepare a chip array, the steps of preparing each pixel are repeated.This can be done automatically with electron beam writing strategy. Thenoble metal reduction is done in parallel when the device is dipped inAgNO₃/HF solution.

FIGS. 8A-8B to FIGS. 10A-10C show SEM images of nanochains. FIG. 11shows an SEM image of a device including a plurality of pixels. FIG. 12shows SEM images of a device including a plurality of pixels and SEMimages of three pixels. FIGS. 13A-13B show a wild type and a mutatedgene BRCA1 in breast cancer. FIGS. 14A-14B show Raman spectra of W1837peptide on a device including nanochains. FIGS. 15A-15B shows Ramanspectra of a mixture of 22 peptides on a device including nanochains.

Other embodiments are within the scope of the following claims.

What is claimed is:
 1. A device for detecting an analyte in a sample comprising: an array including a plurality of pixels, each pixel including a nanochain comprising: a first nanostructure, a second nanostructure, and a third nanostructure, wherein size of the first nanostructure is larger than that of the second nanostructure, and size of the second nanostructure is larger than that of the third nanostructure, and wherein the first nanostructure, the second nanostructure, and the third nanostructure are positioned on a substrate such that when the nanochain is excited by an energy, an optical field between the second nanostructure and the third nanostructure is stronger than an optical field between the first nanostructure and the second nanostructure, wherein the array is configured to receive a sample; and a detector arranged to collect spectral data from a plurality of pixels of the array.
 2. The device of claim 1, wherein the first nanostructure includes a metal, the second nanostructure includes a metal, and the third nanostructure includes a metal.
 3. The device of claim 1, wherein the nanostructure includes silver, gold, platinum, or aluminum.
 4. The device of claim 1, wherein the second nanostructure is positioned between the first nanostructure and the third nanostructure.
 5. The device of claim 1, wherein the distance between the first nanostructure and the second nanostructure is less than the diameter of the second nanostructure.
 6. The device of claim 1, wherein the distance between the second nanostructure and the third nanostructure is less than the diameter of the third nanostructure.
 7. The device of claim 1, wherein the device includes 10 by 10 pixels.
 8. The device of claim 1, wherein the distance between two pixels is at least 1 μm.
 9. The device of claim 1, wherein each pixel delivers spectral data for at least one analyte.
 10. The device of claim 1, wherein each pixel is capable of detecting at least two different kinds of analytes.
 11. The device of claim 1, wherein the device is capable of detecting at least 20 different kinds of analytes.
 12. The device of claim 1, wherein the analyte is detectable by Raman spectrum.
 13. The device of claim 1, wherein the analyte is a peptide.
 14. The device of claim 1, wherein the device is capable of detecting a single molecule.
 15. The device of claim 1, wherein the first nanostructure is a nanosphere, the second nanostructure is a nanosphere, and the third nanostructure is a nanosphere.
 16. A method for detecting an analyte comprising: contacting an analyte with an array including a plurality of pixels, each pixel including a nanochain comprising: a first nanostructure, a second nanostructure, and a third nanostructure, wherein size of the first nanostructure is larger than that of the second nanostructure, and size of the second nanostructure is larger than that of the third nanostructure, and wherein the first nanostructure, the second nanostructure, and the third nanostructure are positioned on a substrate such that when the nanochain is excited by an energy, an optical field between the second nanostructure and the third nanostructure is stronger than the optical field between the first nanostructure and the second nanostructure; and collecting spectral data from a plurality of pixels of the array.
 17. The method of claim 16, further comprising taking Raman spectrum of the nanochain.
 18. The method of claim 16, wherein the analyte includes a peptide.
 19. The method of claim 16, wherein the first nanostructure includes a metal, the second nanostructure includes a metal, and the third nanostructure includes a metal.
 20. The method of claim 16, further comprising detecting a single molecule. 